Multiphoton Imaging of Collagen, Elastin, and Calcification in Intact Soft‐Tissue Samples

Gade, Piyusha S.; Robertson, Anne M.; Chuang, Chih-Yuan

doi:10.1002/cpcy.51

Cited by 16 publications

(6 citation statements)

References 41 publications

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“…Samples were excited at a wavelength of 780 nm, with a constant laser power of 1.35 mW and a depth step-size of 2 μm. These methods are similar to previous reports [ [93] , [94] , [95] ].…”

Section: Methodssupporting

confidence: 89%

Adipose-derived stromal cell secreted factors induce the elastogenesis cascade within 3D aortic smooth muscle cell constructs

Ramaswamy

Sides

Cunnane

et al. 2019

Matrix Biology Plus

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Objective Elastogenesis within the medial layer of the aortic wall involves a cascade of events orchestrated primarily by smooth muscle cells, including transcription of elastin and a cadre of elastin chaperone matricellular proteins, deposition and cross-linking of tropoelastin coacervates, and maturation of extracellular matrix fiber structures to form mechanically competent vascular tissue. Elastic fiber disruption is associated with aortic aneurysm; in aneurysmal disease a thin and weakened wall leads to a high risk of rupture if left untreated, and non-surgical treatments for small aortic aneurysms are currently limited. This study analyzed the effect of adipose-derived stromal cell secreted factors on each step of the smooth muscle cell elastogenesis cascade within a three-dimensional fibrin gel culture platform. Approach and results We demonstrate that adipose-derived stromal cell secreted factors induce an increase in smooth muscle cell transcription of tropoelastin, fibrillin-1, and chaperone proteins fibulin-5, lysyl oxidase, and lysyl oxidase-like 1, formation of extracellular elastic fibers, insoluble elastin and collagen protein fractions in dynamically-active 30-day constructs, and a mechanically competent matrix after 30 days in culture. Conclusion Our results reveal a potential avenue for an elastin-targeted small aortic aneurysm therapeutic, acting as a supplement to the currently employed passive monitoring strategy. Additionally, the elastogenesis analysis workflow explored here could guide future mechanistic studies of elastin formation, which in turn could lead to new non-surgical treatment strategies.

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Section: Methodssupporting

confidence: 89%

Adipose-derived stromal cell secreted factors induce the elastogenesis cascade within 3D aortic smooth muscle cell constructs

Ramaswamy

Sides

Cunnane

et al. 2019

Matrix Biology Plus

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“…1E) and detected no signals classified as GFP-Salmonella in the spleen of mice infected with yellow fluorescent protein (YFP)-Salmonella indicating specific detection. Collagen second-harmonic signals (54) and in vivo staining of host cell surface antigens with fluorescence-labeled antibodies revealed the tissue context of Salmonella cells (Fig. 1 D and E).…”

Section: Resultsmentioning

confidence: 98%

Tissue compartmentalization enablesSalmonellapersistence during chemotherapy

Claudi

Fanous

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

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Antimicrobial chemotherapy can fail to eradicate the pathogen, even in the absence of antimicrobial resistance. Persisting pathogens can subsequently cause relapsing diseases. In vitro studies suggest various mechanisms of antibiotic persistence, but their in vivo relevance remains unclear because of the difficulty of studying scarce pathogen survivors in complex host tissues. Here, we localized and characterized rare surviving Salmonella in mouse spleen using high-resolution whole-organ tomography. Chemotherapy cleared >99.5% of the Salmonella but was inefficient against a small Salmonella subset in the white pulp. Previous models could not explain these findings: drug exposure was adequate, Salmonella continued to replicate, and host stresses induced only limited Salmonella drug tolerance. Instead, antimicrobial clearance required support of Salmonella-killing neutrophils and monocytes, and the density of such cells was lower in the white pulp than in other spleen compartments containing higher Salmonella loads. Neutrophil densities declined further during treatment in response to receding Salmonella loads, resulting in insufficient support for Salmonella clearance from the white pulp and eradication failure. However, adjunctive therapies sustaining inflammatory support enabled effective clearance. These results identify uneven Salmonella tissue colonization and spatiotemporal inflammation dynamics as main causes of Salmonella persistence and establish a powerful approach to investigate scarce but impactful pathogen subsets in complex host environments.

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“…For this matter, it is important to filter the image and to recover an accurate representation of the fiber network through segmentation (Hompland et al 2008) or fiber extraction (Wegner et al 2017). Gade et al (2019) performed a segmentation on the SHG stack using the Otsu thresholding. Then, the authors computed the area of segmented pixels in every slice and sum up the segmented area across the volume to calculate total areal density in the image stack.…”

Section: Scale Of Measurementioning

confidence: 99%

“…In general, SHG images are segmented separately. For example, in Gade et al (2019) and Balu et al (2014), the authors segmented the SHG images using a thresholding technique. Then, they calculated the number of white pixels in every image and summed them up across the volume to calculate total 3D density.…”

Section: Input Data Nature (2d/projected 3d/3d)mentioning

confidence: 99%

Quantitative analysis of second harmonic generated images of collagen fibers: a review

et al. 2022

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Purpose The human body is a complex structure. Its strength is ensured by the collagen protein which exists under the form of fibers. The quantitative analysis of these fibers in biological tissues can be very interesting to establish a relationship between the microstructure and their functions. This analysis is usually performed using two-photon microscopy and second harmonic generated (SHG) images. Lately, more and more researchers focused on the use of SHG images since it is a non-invasive technique and allows the capture of collagen fibers only. Many image-processing techniques can be used to extract quantitative information from those images such as fiber orientations, dimensions, and density. Therefore, accurate measure extraction depends mainly on the used image processing methods and, thus, it is necessary to know what processing technique to use. Methods The main purpose of this article is to exhibit the most used techniques in collagen fiber quantitative analysis then categorize them according to the information to extract. A comparison of three most used methods in fiber orientation’s estimation is carried out. Result and conclusion Despite the considerable number of papers aiming to quantitatively analyze collagen fibers from SHG images, two main aspects were not deeply covered. First, the use of deep learning algorithms is still limited even for segmentation and denoizing applications. Second, most of the studies processed in this review focused on two-dimensional SHG images and did not take into consideration collagen fibers as a three-dimensional volume.

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Multiphoton Imaging of Collagen, Elastin, and Calcification in Intact Soft‐Tissue Samples

Cited by 16 publications

References 41 publications

Adipose-derived stromal cell secreted factors induce the elastogenesis cascade within 3D aortic smooth muscle cell constructs

Adipose-derived stromal cell secreted factors induce the elastogenesis cascade within 3D aortic smooth muscle cell constructs

Tissue compartmentalization enablesSalmonellapersistence during chemotherapy

Quantitative analysis of second harmonic generated images of collagen fibers: a review

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